Establishment of Hairy Root Transformation System of Melilotus albus Induced by Agrobacterium rhizogenes

doi:10.11733/j.issn.1007-0435.2021.11.026

Acta Agrestia Sinica ›› 2021, Vol. 29 ›› Issue (11): 2591-2599.DOI: 10.11733/j.issn.1007-0435.2021.11.026

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Establishment of Hairy Root Transformation System of Melilotus albus Induced by Agrobacterium rhizogenes

WANG Sheng-sheng, DUAN Zhen, ZHANG Ji-yu

State Key Laboratory of Grassland Agro-Ecosystems, College of Pastoral Agriculture Science and Technology, Lanzhou University, Lanzhou, Gansu Province 730020, China

Received:2021-06-07 Revised:2021-08-16 Online:2021-11-15 Published:2021-12-06

发根农杆菌介导的白花草木樨毛状根转化体系的建立

王升升, 段珍, 张吉宇

兰州大学草地农业生态系统国家重点实验室, 兰州大学草地农业科技学院, 甘肃兰州 730020

通讯作者: 张吉宇,E-mail:zhangjy@lzu.edu.cn
作者简介:王升升(1996-),女,汉族,甘肃天水人,硕士研究生,主要从事草类作物遗传育种研究,E-mail:wangshsh19@lzu.edu.cn
基金资助:
国家自然科学基金-国际（地区）合作与交流项目（NSFC-CGIAR）（32061143035）和甘肃省科技重大专项（19ZD2NA002-2）资助

Abstract

Abstract: Melilotus albus is one of the most common cultivated species of M. spp,which has high resistance and nitrogen fixation ability. In this study,we established an efficient hairy root transformation system induced by Agrobacterium rhizogenes in M. albus,and the nodulation gene MaROP6 was used as the target gene. The results showed that hairy roots were obtained within two weeks with a rate of 78%,and the positive transgenic hairy roots of MaROP6 gene reached 70.8%. The qRT-PCR results showed that the average relative expression of MaROP6 in transgenic hairy roots was 36.87 times higher than that in the control. The establishment of hairy root transformation system laid a foundation for identifying the gene function of M. albus.

Key words: Melilotus albus, Agrobacterium rhizogenes, Hairy root transformation, Gene expression

摘要： 白花草木樨（Melilotus albus Medic.ex Desr.）是草木樨最常见的栽培用种之一，具有较强的抗逆性和固氮能力。为了开发一套高效的草木樨转化体系，本研究以白花草木樨为材料，以结瘤基因MaROP6为目的基因，建立了发根农杆菌（Agrobacterium rhizogenes）介导的高效白花草木樨毛状根转化体系。结果表明：利用该方法可以在两周左右获得毛状根，其发生率可达78%，MaROP6基因的毛状根转化率可达70.8%。qRT-PCR鉴定结果表明，MaROP6转基因毛状根的平均相对表达量为对照的36.87倍。毛状根转化体系的建立为白花草木樨基因功能的验证奠定了基础。

关键词: 白花草木樨, 发根农杆菌, 毛状根转化, 基因表达

CLC Number:

S551.6

WANG Sheng-sheng, DUAN Zhen, ZHANG Ji-yu. Establishment of Hairy Root Transformation System of Melilotus albus Induced by Agrobacterium rhizogenes[J]. Acta Agrestia Sinica, 2021, 29(11): 2591-2599.

王升升, 段珍, 张吉宇. 发根农杆菌介导的白花草木樨毛状根转化体系的建立[J]. 草地学报, 2021, 29(11): 2591-2599.

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References

[1] WU F,LUO K,YAN Z Z,et al. Analysis of miRNAs and their target genes in five Melilotus albus NILs with different coumarin content[J]. Scientific Reports,2018,8(1):1-13
[2] BRENNER D M. Methods for Melilotus germplasm regeneration[J]. Plant Genetic Resources Newsletter,2005(141):51-55
[3] 景春梅,刘慧,席琳乔,等. 优质牧草、绿肥草木樨的研究进展[J]. 草业科学,2014,31(12):2308-2315
[4] LUO K,JAHUFER M Z Z,ZHAO H,et al. Genetic improvement of key agronomic traits in Melilotus albus[J]. Crop Science,2018(58):1-10
[5] 陈立强,白小明,蔺胜权. 阿拉尔野生一年生草木樨资源的生产性能和营养品质研究[J]. 草地学报,2020,28(6):1643-1649
[6] HODGSON H J,BULA R J. Hardening behavior of sweetclover (Melilotus Spp.) varieties in a subarctic environment[J]. Agronomy Journal,1956,48(4):157-160
[7] 李国文. 草木樨的饲用价值和栽培管理技术[R]. 现代畜牧科技,2017,3(38):44-44
[8] 格根图,刘燕,贾玉山. 草木樨干草营养价值及饲喂绒山羊的效果研究[J]. 草地学报,2013,21(2):401-405
[9] ROZEMA B J. Symbiotic nitrogen fixation in legumes:Perspectives for saline agriculture[J]. Environmental Experimental Botany,2013(92):134-143
[10] CONN J S,SEEFELDT S S. Invasive white sweetclover (Melilotus officinalis) control with herbicides,cutting,and flaming[J]. Invasive Plant Science and Management,2009,2(3):270-277
[11] QUIJANO-CELIS C E,PINO J A,MORALES G. Chemical composition of the leaves essential oil of Melilotus officinalis (L.) pallas from colombia[J]. Journal of Essential Oil Bearing Plants,2010,13(3):313-315
[12] YAN Z Z,WU F,LUO K,et al. Cross-species transferability of EST-SSR markers developed from the transcriptome of Melilotus and their application to population genetics research[J]. Scientific Reports,2017,7(8):1-11
[13] WU F,ZHANG D Y,MA J X,et al. Analysis of genetic diversity and population structure in accessions of the genus Melilotus[J]. Industrial Crops Products,2016(85):84-92
[14] 骆凯,狄红艳,张吉宇,等. 19份草木樨种质农艺学与品质性状初步评价[J]. 草业科学,2014,31(11):2125-2134
[15] DI H Y,DUAN Z,LUO K,et al. Interspecific phylogenic relationships within genus Melilotus Based on nuclear and chloroplast DNA[J]. Plos One,2015,10(7):1-12
[16] 剡转转,任艳,吴凡,等. 白花草木樨EST-SSR标记的开发与筛选[J]. 草业科学,2017,34(9):1802-1814
[17] 狄红艳,骆凯,张吉宇,等. 基于ITS和trnL-trnF序列的草木樨种群遗传多样性研究[J]. 西北植物学报,2014,34(2):265-269
[18] HIRSCH A M,LEE A,DENG W M,et al. An open-flower mutant of Melilotus alba:Potential for floral-dip transformation of a papilionoid legume with a short life cycle[J]. American Journal of Botany,2010,97(3):395-404
[19] LUO K,WU F,ZHANG D Y,et al. Transcriptomic profiling of Melilotus albus near-isogenic lines contrasting for coumarin content[J]. Scientific Reports,2017,7(1):1-10
[20] 邵明成,王大鹏. 发根农杆菌诱导产生发状根及其在植物科学领域中的应用[J]. 黑龙江农业科学,2015(3):146-150
[21] BOISSON D A. Agrobacterium rhizogenes-transformed roots of Medicago truncatula for the study of nitrogen-fixing and endomycorrhizal symbiotic associations[J]. Molecular Plant-Microbe Interactions,2001,14(6):695-700
[22] TOTH K,BATEK J,STACEY G. Generation of soybean (Glycine max) transient transgenic roots[J]. Current Protocols in Plant Biology,2016,1(1):1-13
[23] KE D X,FANG Q,CHEN C F,et al. The Small GTPase ROP6 interacts with NFR5 and is involved in nodule formation in Lotus japonicus[J]. Plant Physiology,2012,159(1):131-143
[24] LIU W,CHEN A M,LUO L, et al. Characterization and expression analysis of Medicago truncatula ROP GTPase family during the early stage of symbiosis[J]. Journal of Integrative Plant Biology,2010,52(7):639-652
[25] 陈安乐. 大豆发根转化方法的建立及GmFRD3在大豆耐铝性中的作用[D]. 长春:吉林大学,2014:26-55
[26] 张吉宇,王彦荣,南志标. 相对定量和绝对定量-以CsSAMDC基因表达分析为例[J]. 中国生物工程杂志,2009,29(8):86-91
[27] ZHENG Z L,YANG Z. The Rop GTPase:an emerging signaling switch in plants[J]. Plant Molecular Biology,2000,44(1):1-9
[28] 陈秀清. 发根农杆菌诱导毛状根研究进展[J]. 安徽农业科学,2011,341(16):9512-9514
[29] 刘艳菊. 发根农杆菌介导的Bt基因Cry8e转化大豆抗蛴螬研究[D].河南:河南师范大学,2010:7-9
[30] 陈燕,李聪,苏加楷,等. 发根农杆菌介导的百脉根的基因转化[J]. 草地学报,1996,4(2):116-120
[31] 王天佐,张文浩. 发根农杆菌介导的花苜蓿毛状根转化体系的建立[J]. 草地学报,2020,28(1):268-272
[32] 刘佳. 苦参离体培养及发根农杆菌诱导其毛状根的研究[D]. 山西:山西农业大学,2015:35-36
[33] 熊笙屹,厍润祥,张璐,等. 转基因植物发根农杆菌研究的进展及应用. 农业与技术,2017,37(16):72-74
[34] 刘伟,郝建平. 发根农杆菌的研究进展及其应用[J]. 山西农业科学,2007,35(7):13-16
[35] 梅错. 箭筈豌豆毛状根转化体系的建立及其验证[D]. 兰州:兰州大学,2020:41-55,196-1209
[36] SI Z Y,YANG Q Q,LIANG R G,et al. Digalactosyldiacylglycerol synthase gene MtDGD1 plays an essential role in nodule development and nitrogen fixation[J]. Molecular plant-microbe interactions,2019,32(9):1
[37] 宗晓秋. 发根农杆菌诱导大豆毛状根体系的建立与应用[D]. 北京:中国农业科学院,2012:38-39
[38] 徐洪伟. 发根农杆菌诱导玉米毛状根发生及再生植株[D]. 长春:东北师范大学,2007:23-33
[39] 任如意,薛巨坤,国会艳,等. 北玄参毛状根诱导及其植株再生[J]. 植物学报,2017,52(6):783-787

Establishment of Hairy Root Transformation System of Melilotus albus Induced by Agrobacterium rhizogenes

发根农杆菌介导的白花草木樨毛状根转化体系的建立

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